Elevated activity of membrane receptor and soluble protein tyrosine kinases (PTKs), occurs in a significant portion of breast tumors, and is indicative of a poor prognosis. These enzymes play a critical role in the regulation of normal cell growth and differentiation; their overexpression may confer a growth advantage to breast cancer cells by increasing sensitivity to locally acting peptide growth factors. It is important to define the role of less well-characterized PTKs as novel components of known signaling pathways also overexpressed in a large proportion of human breast cancers. A novel non-receptor PTK (Brk) was cloned from a human metastatic breast tumor, and found to be overexpressed in human breast carcinomas and breast cancer cell lines, but not in normal tissue. Although clearly in a novel family of soluble PTKs, Brk most closely resembles the Src oncogene, containing one SH3- and one SH2-domain. Overexpression of Brk transformed human mammary epithelial cells. This protein is associated with the epidermal growth factor (EGF) receptor in human breast cancer cells. Preliminary data suggests that EGF and progestins synergistically regulate Brk. It is hypothesized that Brk confers a growth advantage to human breast cancer cells by acting as a progestin-regulated signaling component in the EGF pathway. The hormonal control of Brk regulation will be addressed by : 1) defining Brk activation by upstream signals, 2) identifying regulatory phosphorylation sites of Brk in response to these signals, and 3) defining signaling pathways downstream of Brk by classical strategies involving "pull-down" co-immunoprecipitation assays. The goal of the proposed studies is to demonstrate a role for signal transduction by a novel PTK (Brk) in breast cancer by defining the regulation and function of Brk as influenced by ovarian steroid hormones and peptide growth factors relevant to breast cancer cell biology.